In mammals, the sensation of pain triggered by thermal, mechanical or chemical stimuli is a useful warning and protective system. Considerable efforts have been put into elucidating the biochemical mechanisms involved in the detection, transduction and transmission of hot and cold sensations in neuronal tissues. Thermal stimuli activate specialized receptors located on sensory neurons, such as those deriving from the dorsal root ganglion (DRG) and the trigeminal ganglion (TG). When these stimuli are in the noxious range (i.e., very hot or cold), they activate a certain subset of thermal receptors on a sub-population of sensory neurons called nociceptors (pain-sensing neurons). Upon activation, the thermal receptors (e.g., ion channels) transduce the noxious stimulus into an electrical signal that is propagated along the sensory neuron to the spinal cord, where it is relayed to the brain, ultimately leading to the perception of pain. Accordingly, these thermal receptors represent highly promising targets for developing drugs for the treatment of various painful conditions.
Several temperature-activated receptors have been identified with wide ranging temperature sensitivities from noxious heat to noxious cold. These temperature-activated receptors belong to the transient receptor potential (TRP) family of non-selective cation channels, which in C. elegans and D. melanogaster are involved in mechano- and osmoregulation. Several of these temperature-activated receptors, including TRPV1 and TRPV2, are implicated in noxious heat sensation (Caterina et al., 1997, Nature, 389: 816; and Caterina et al., 1999, Nature 398: 436). TRPV1, the most extensively characterized member of the thermo-TRP family, is activated by moderate heat (˜43° C.), capsaicin, protons and certain endocannabinoids, such as anandamide and 2-AG. It is well accepted that TRPV1 contributes to acute thermal nociception and hyperalgesia after injury (Clapham, Nature. 2003, 426(6966): 517-24).
TRPV2, also termed VRL-1, has been proposed as a sensor of noxious temperatures (>52° C.), which presumably mediates “first” pain, i.e. the rapid, acute, and sharp pain evoked by noxious stimuli (Caterina et al., 1999, supra; Story et al., Cell, 2003, 112:819-829, and references therein). TRPV2 is structurally most closely related to TRPV1 (˜50% sequence identity at the protein level). TRPV2 is expressed in medium- to large- diameter neurons of sensory ganglia, as well as at lower levels in brain, spinal cord, spleen and lung. Furthermore, TRPV2 is upregulated in sympathetic postganglionic neurons following injury, suggesting a potential role for TRPV2 in sympathetically mediated pain (Gaudet et al., Brain Res. 2004, 1017(1-2):155-62). Thus, modulation of TRPV2 may potentially have many therapeutic applications.
Despite great interest in TRPV2 modulation, a system for screening, identifying and characterizing TRPV2 modulators has yet to be developed. This is in part due to the lack of known, and in particular, selective TRPV2 agonists, as well as the technical difficulty of assaying these channels in a high temperature environment. In general, TRPV2 does not respond to known TRPV1 agonists (Benham et al., 2003, Cell Calcium 33:479-487). However, a recent study reported that 2-aminoethoxydiphenyl borate (2-APB), a non-selective TRP modulator, was able to activate TRPV1, TRPV2, and TRPV3 (Hu et al (2004), J. Biol. Chem., 279: 35741-8), although TRPV2 activation by 2-APB was not observed by others (Chung et al. (2004), J. Neurosci. 24: 5177-82).
In an effort to overcome the above-mentioned challenges, the present invention provides novel compositions and methods for screening, identifying and characterizing TRPV2 agonists.